There has been much interest in the past few years regarding the role of nitric oxide (NO) in a variety of physiological phenomenon, such as the control of blood flow and vasodilation as well as participating in bacteriocidal and tumoricidal activity. A number of different compounds have been used to deliver nitric oxide to biological media. We previously showed that the NONOate complexes sensitize hypoxic cells to ionizing radiation using in vitro model systems while in vivo NO administered to animals provides protection against whole body irradiation. In vitro experiments show that NO can modulate chemotherapeutic agents. The cytotoxicity of alkylating agents, melphalan and thiotepa, are enhanced by NO, while NO protects against cisplatin. Chemotherapeutic studies in vivo using a murine tumor model have shown that the combination of melphalan and NO offers significantly more tumor control than melphalan alone. Since NO has been shown to be a key molecule in regulating vascular tone, it may be important in opening blood flow to hypoxic regions in tumors and reducing interstitial fluid pressure in tumors. We have examined the effects of both S-nitrosothiols and NONOates on interstitial pressure finding little effect. However, nitric oxide synthase inhibitors significantly lower pressure offering a potential strategy for enhancement of drug and oxygen delivery to tumor. We previously demonstrated that NO provides protection against peroxide mediated toxicity in mammalian cells, suggesting a potential therapy against conditions involving oxidative stress injury. Examination of peroxide and superoxide mediated toxicity in bacteria reveals that NO enhances toxicity. This opposite effect from mammalian cells suggests that NO and peroxide are key participants in the immune response to foreign pathogens. In our in vivo and in vitro experiments with NO, we have begun to appreciate timing as a key factor in successfully using NO/NO donor complexes clinically. We are developing a number of different analytical tools to better understand the biological mechanisms of NO and to aid in formulating successful protocols.